The role of position a in determining the stability and oligomerization state of α-helical coiled coils: 20 amino acid stability coefficients in the hydrophobic core of proteins

• Published in: Protein science 1999-11, Vol.8 (11), p.2312-2329
• Main Authors: WAGSCHAL, KURT, TRIPET, BRIAN, LAVIGNE, PIERRE, MANT, COLIN, HODGES, ROBERT S.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Circular Dichroism; Disulfides; Drug Stability; Guanidine; hydrophobic core; Macromolecular Substances; Models, Molecular; Molecular Sequence Data; oligomerization state; Peptides - chemical synthesis; Peptides - chemistry; Protein Denaturation; protein design; protein stability; Protein Structure, Secondary; Proteins - chemistry; Regression Analysis; Thermodynamics; Urea; α‐helical coiled coil
• ISSN: 0961-8368; 1469-896X
• DOI: 10.1110/ps.8.11.2312

We describe here a systematic investigation into the role of position a in the hydrophobic core of a model coiled-coil protein in determining coiled-coil stability and oligomerization state. We employed a model coiled coil that allowed the formation of an extended three-stranded trimeric oligomerization state for some of the analogs; however, due to the presence of a Cys-Gly-Gly linker, unfolding occurred from the same two-stranded monomeric oligomerization state for all of the analogs. Denaturation from a two-stranded state allowed us to measure the relative contribution of 20 different amino acid side chains to coiled-coil stability from chemical denaturation profiles. In addition, the relative hydrophobicity of the substituted amino acid side chains was assessed by reversed-phase high-performance liquid chromatography and found to correlate very highly (R = 0.95) with coiled-coil stability. We also determined the effect of position a in specifying the oligomerization state using ultracentrifugation as well as high-performance size-exclusion chromatography. We found that nine of the analogs populated one oligomerization state exclusively at peptide concentrations of 50 μM under benign buffer conditions. The Leu-, Tyr-, Gln-, and His-substituted analogs were found to be exclusively three-stranded trimers, while the Asn-, Lys-, Orn-, Arg-, and Trp-substituted analogs formed exclusively two-stranded monomers. Modeling results for the Leu-substituted analog showed that a three-stranded oligomerization state is preferred due to increased side-chain burial, while a two-stranded oligomerization state was observed for the Trp analog due to unfavorable cavity formation in the three-stranded state.